The use of optimal cutting temperature compound can inhibit amplification by polymerase chain reaction.
Optimal cutting temperature (OCT) is a widely used embedding medium for tissues for histopathologic analysis. This investigation examined the effects that OCT storage can have upon the ability to perform subsequent molecular biological analyses. Tumor material was dissected into small pieces and stored at approximately 20 degrees C both with and without OCT. DNA and RNA were then extracted from the tissue fragments and analyzed by the polymerase chain reaction (PCR), using primer sets designed to amplify a range of product sizes, and also by reverse transcriptase-PCR (RT-PCR). The storage of pathological specimens in OCT compound was found to affect significantly and irreversibly the ability to amplify DNA in the PCR, particularly as the size of the amplified fragment increased. This effect appeared to occur as a result of greater degradation of DNA extracted from tissue embedded in OCT compared to DNA extracted from tissue stored without OCT. RNA quality appeared unaffected, which may be because of the extraction protocol employed. Our results suggest that OCT-embedded frozen-tissue samples may be used for RNA isolation for subsequent RT-PCR and for the in vitro amplification of DNA targets of approximately < 300 base pairs only. We strongly advise against the routine storage of any tissue biopsy material in OCT if molecular analyses may be required.
Turbett
G R
GR
Department of Pathology, Royal Perth Hospital, Western Australia, Australia.
Sellner
L N
LN
D005629:Frozen Sections; D005819:Genetic Markers; D006801:Humans; D008228:Lymphoma, Non-Hodgkin / Q000235:genetics / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods; D016610:Tissue Embedding / Q000379:methods*
D005819:Genetic Markers